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Lamin-Related Congenital Muscular Dystrophy Alters Mechanical Signaling and Skeletal Muscle Growth

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Lamin-Related Congenital Muscular Dystrophy Alters Mechanical Signaling and Skeletal Muscle Growth International Journal of Molecular Sciences Article Lamin-Related Congenital Muscular Dystrophy Alters Mechanical Signaling and Skeletal Muscle Growth Daniel J. Owens 1,2 , Julien Messéant 1, Sophie Moog 3, Mark Viggars 2, Arnaud Ferry 1,4, Kamel Mamchaoui 1,5, Emmanuelle Lacène 5 , Norma Roméro 5,6, Astrid Brull 1 , Gisèle Bonne 1 , Gillian Butler-Browne 1 and Catherine Coirault 1,* 1 Center for Research in Myology, Sorbonne Université, INSERM UMRS_974, 75013 Paris, France; [email protected] (D.J.O.); [email protected] (J.M.); [email protected] (A.F.); [email protected] (K.M.); [email protected] (A.B.); [email protected] (G.B.); [email protected] (G.B.-B.) 2 Research Institute for Sport and Exercise Science, Liverpool John Moores University, Liverpool L3 3AF, UK; [email protected] 3 Inovarion, 75005 Paris, France; [email protected] 4 Université de Paris, 75006 Paris, France 5 Neuromuscular Morphology Unit, Institute of Myology, Pitié-Salpêtrière Hospital, 75013 Paris, France; [email protected] (E.L.); [email protected] (N.R.) 6 APHP, Reference Center for Neuromuscular Disorders, Pitié-Salpêtrière Hospital, Institute of Myology, 75013 Paris, France * Correspondence: [email protected]; Tel.: +33-1-1-4216-5708 Abstract: Laminopathies are a clinically heterogeneous group of disorders caused by mutations in the LMNA gene, which encodes the nuclear envelope proteins lamins A and C. The most frequent diseases associated with LMNA mutations are characterized by skeletal and cardiac involvement, and include autosomal dominant Emery–Dreifuss muscular dystrophy (EDMD), limb-girdle muscular Citation: Owens, D.J.; Messéant, J.; dystrophy type 1B, and LMNA-related congenital muscular dystrophy (LMNA-CMD). Although Moog, S.; Viggars, M.; Ferry, A.; the exact pathophysiological mechanisms responsible for LMNA-CMD are not yet understood, Mamchaoui, K.; Lacène, E.; Roméro, severe contracture and muscle atrophy suggest that mutations may impair skeletal muscle growth. N.; Brull, A.; Bonne, G.; et al. Using human muscle stem cells (MuSCs) carrying LMNA-CMD mutations, we observe impaired Lamin-Related Congenital Muscular myogenic fusion with disorganized cadherin/β catenin adhesion complexes. We show that skeletal Dystrophy Alters Mechanical Signaling muscle from Lmna-CMD mice is unable to hypertrophy in response to functional overload, due and Skeletal Muscle Growth. Int. J. to defective fusion of activated MuSCs, defective protein synthesis and defective remodeling of Mol. Sci. 2021, 22, 306. https:// the neuromuscular junction. Moreover, stretched myotubes and overloaded muscle fibers with doi.org/10.3390/ijms22010306 LMNA-CMD mutations display aberrant mechanical regulation of the yes-associated protein (YAP). We also observe defects in MuSC activation and YAP signaling in muscle biopsies from LMNA-CMD Received: 22 October 2020 patients. These phenotypes are not recapitulated in closely related but less severe EDMD models. Accepted: 26 December 2020 In conclusion, combining studies in vitro, in vivo, and patient samples, we find that LMNA-CMD Published: 30 December 2020 mutations interfere with mechanosignaling pathways in skeletal muscle, implicating A-type lamins Publisher’s Note: MDPI stays neu- in the regulation of skeletal muscle growth. tral with regard to jurisdictional clai- ms in published maps and institutio- Keywords: mechanotransduction; muscle growth; nuclear envelope; satellite cell; YAP nal affiliations. 1. Introduction Copyright: © 2020 by the authors. Li- censee MDPI, Basel, Switzerland. Skeletal muscle is a highly organized tissue designed to produce force and movement. This article is an open access article It is largely composed of differentiated, multinucleated, postmitotic myofibers responsi- distributed under the terms and con- ble for contraction, and also contains a population of mononucleated muscle stem cells ditions of the Creative Commons At- (MuSCs), called satellite cells, that reside between myofibers and the surrounding basal tribution (CC BY) license (https:// lamina and that display long-term quiescence. Following muscle injury, during postnatal creativecommons.org/licenses/by/ growth and in response to many hypertrophic responses, MuSCs are activated and un- 4.0/). dergo a highly orchestrated series of events that regulate their proliferation, polarity, and Int. J. Mol. Sci. 2021, 22, 306. https://doi.org/10.3390/ijms22010306 https://www.mdpi.com/journal/ijms Int. J. Mol. Sci. 2021, 22, 306 2 of 21 differentiation (reviewed in [1]). Although a subset of MuSCs return to quiescence [2], other activated MuSCs subsequently differentiate and fuse to each other or to existing my- ofibers. Adhesive contacts between activated MuSCs or between MuSCs and the myofibers are critical to sense and transduce intracellular forces between cells and the extracellular matrix [3,4] and neighboring cells [5–7], and provide direct signaling cues essential to stem cell behavior [8]. Apart from cell adhesive components, recent studies clearly establish that the nucleus is critical for cells to sense and respond to the mechanical properties of their environment [9,10], thus implicating that muscle plasticity depends on nuclear mechanotransduction. The mechanical properties of the nucleus are largely determined by the nuclear lamina, a fibrous meshwork composed of lamin intermediate filament proteins that underlies the inner nuclear membrane. Nuclear lamins are encoded by three genes; lamin-A and lamin-C (known as A-type lamins) are alternatively spliced products of the LMNA gene, whereas lamin-B1 and lamin-B2 (B-type lamins) are encoded by the LMNB1 and LMNB2 genes. Mutations in the LMNA gene cause laminopathies, a phenotypically diverse group of disorders, including muscular dystrophies and cardiomyopathies [11]. The majority of LMNA mutations cause the autosomal dominant Emery–Dreifuss muscular dystrophy or EDMD, characterized by progressive muscle wasting, contractures, and cardiomyopathy. Lamin-related congenital muscular dystrophy (LMNA-CMD) manifests as a particularly severe skeletal muscle phenotype, with muscle wasting beginning very early in life [12], frequent nuclear defects [13], and impaired mechanosensing [14]. Cellular mechanotransduction involves physical connection of the nuclear lamina to the cytoskeleton [15]. Such connections are mediated by the members of nucleoskeleton and cytoskeleton (LINC complex) [16] that comprise SUN domain proteins that bind via their nucleoplasmic domains to A-type lamins [16] and nesprins at the outer nuclear membrane that bind to the cytoskeleton [17]. Together A-type lamins and LINC complexes are crucial for mechanical coupling between the nucleoskeleton and the cytoskeleton [10,15]. Functional loss in A-type lamins alters cytoskeletal actin structures around the nucleus in cells cultured on a rigid substrate [18–20], presumably through an impaired activation of the mechanosensitive transcriptional cofactor serum responsive factor (SRF) and its target genes [21]. LMNA-CMD mutations also compromise the ability of cells to adapt their actin cytoskeleton to different cellular microenvironments and to withstand mechanical stretching of the extracellular matrix, owing to the deregulation of yes-associated protein (YAP) [14], a cotranscriptional factor that nuclear or cytoplasmic localization is modulated by diverse biomechanical signals from the actin cytoskeleton [22]. Collectively, these results implicate A-type lamins in modulating the dynamics and organization of the actin cytoskeleton and thus are also involved in cellular mechanotransduction. It is currently unknown whether mechanotransduction defects in LMNA-CMD muta- tions may explain abnormal skeletal muscle growth seen in laminopathic patients. In the current study, we aim to investigate the role of A-type lamins in the regulation of mechan- otransduction at cell–cell adhesions and in multinucleated muscle cells. We analyzed three different human cell lines with LMNA mutations responsible for congenital muscle dystrophy, namely, LMNA c.94_96delAAG, LMNA p.Arg249Trp, and LMNA p.Leu380Ser. All these mutations are localized in the head (LMNA c.94_96delAAG) or in the rod do- main (p.Arg249Trp and p.Leu380Ser) of the A-type lamin and are predicted to modify the oligomerization state of the proteins. In other words, these mutations affect the structure and integrity of the nucleoskeleton ([23], compromise the mechanical properties of the nucleus, and force transmission between the nucleoskeleton and the cytoskeleton [15,23]. We also want to determine the consequences of A-type lamin mutations on in vivo muscle adaptation to a mechanical challenge. We hypothesize that LMNA-CMD mutations im- pair cellular and molecular mechanisms contributing to skeletal muscle growth. For the first time, we show that LMNA-CMD mutations impaired myogenic fusion in vitro due to disorganized cadherin/β-catenin complexes with reduced M-cadherin and β-catenin protein expression. Defective skeletal muscle growth was also revealed in vivo, since Int. J. Mol. Sci. 2021, 22, 306 3 of 21 the Lmna-CMD mouse model was unable to hypertrophy due to defective accretion of activated satellite cells in response to functional overload. Moreover, myotubes and muscle fibers with LMNA-CMD mutations demonstrate aberrant regulation of YAP nucleocyto- plasmic translocation in response to different mechanical challenges, which may explain the reduced protein
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